Selective detection schemes are of great importance in determining targeted components from complex matrices. This strategy is useful when studying the disposition of drugs, as is necessary for their commercial release and to understand their mechanisms of action or toxicity, or when evaluating fundamental biochemical processes which underlie studies of health and disease. With continued improvements over recent years, the chemical reaction interface/mass spectrometer (CRIMS) approach that was developed by the P.I. has emerged as one of the most powerful methods devised for carrying out these searches when the targets have some unique isotopic or elemental signature. CRIMS can selectively detect 13C, 15N, and 2H labeled drugs from biological specimens at concentrations at or below l ng/ml. Molecules containing just one atom of sulfur or chlorine have limits below 100 pg on-column. These analyses are linear over 3-4 orders of magnitude. Recent experiments show that fluorine-based CRIMS chemistry is quite powerful, in particular for phosphorus-selective detection. Isotope ratios can be quantified at levels challenged only by special isotope-ratio mass spectrometers. Recently, the coupling of an HPLC and CRIMS produced an instrumental configuration that offers remarkable power and versatility. Its potential as a universal detector will continue to be evaluated. With more effective use of stable isotopes, CRIMS should reduce the number of experiments where radioactivity is used. The focus of this proposal is to continue to improve upon and expand the current capabilities of this still-emerging method. The goals of this application contain a combination of: chemical innovation; improved chromatography and instrumentation; and new analytical dimensions for CRIMS. HPLC separations, increasingly important in biomedical analyses, can benefit particularly from CRIMS because of the limited range of selective detection schemes for HPLC. Chemical innovations should expand and improve the selective detection capabilities of CRIMS. To evaluate how well CRIMS can assist in biomedical analyses, a selected group of applications are proposed using either isotopic or elemental labels. A particular focus for such studies involves macromolecules and HPLC/CRIMS. The use of highly and uniformly stable-isotope labeled proteins should allow unprecedented capabilities in studying their metabolism. For drugs or other xenobiotics that contain a target element, such as chlorine, covalent binding to macromolecules should be detected without digesting them.